Retinal development, normal function, and disease are controlled, to a large extent, by the pattern of genes expressed by the cells of the retina. Despite the significant advances being made in understanding the role of epigenetics in gene regulation in other fields, little is known about the relationship between DNA methylation patterns, retinal gene expression, and retinal disease. In the United States alone, almost 4 million individuals currently have either glaucoma or age-related macular degeneration (AMD), and untreatable blindness can result despite treatment. With both diseases, gene expression changes in the retina have been observed. One modulator of gene expression is DNA methylation. We hypothesize that alterations in DNA methylation, accompany and may actually precede the gene expression changes seen with the onset of glaucoma and AMD. The proposed research will explore this hypothesis by providing a detailed, genome-wide map of the DNA methylation changes in the human retina that are associated with these two diseases. Disease-affected cell populations (retinal ganglion cells in glaucoma;photoreceptors and retinal pigment epithelium in AMD) will be individually isolated by laser capture microdissection, and compared to the same cell population from age- and sex-matched normal control eyes. Genomic DNA samples will be enriched for methylated DNA by affinity purification using immobilized MBD2-MBD. Methylated-enriched samples will be hybridized to high resolution human tiling microarray sets. Unenriched genomic DNA will be separately hybridized for calculations to adjust for probe effects. Differentially methylated areas will be identified by pair- wise comparisons between normal and diseased eyes. Nearby annotated genes will be identified as candidates for having disease-specific DNA methylation patterns and will be further investigated. The ability of epigenetic mechanisms, such as DNA methylation, to modulate patterns of gene expression in the setting of eye disease offers an entirely new frontier for exploration and has the potential to significantly affect our understanding and treatment of important ocular diseases such as glaucoma and AMD. Public Health Relevance: The ability of epigenetic mechanisms, such as DNA methylation, to initiate and modulate abnormal gene expression in the retina offers an entirely new frontier for investigation. Exploring the role of DNA methylation in the etiology, severity, and progression of glaucoma and age-related macular degeneration should lead to novel insights into the molecular mechanisms of disease pathogenesis and innovative therapeutic approaches.